长链非编码RNA AC005062.1通过调控结肠癌转移相关基因1的表达对结直肠癌恶性表型的影响

lncRNA AC005062.1 affects the malignant phenotype of colorectal cancer by regulating the expression of MACC1

目的探究长链非编码RNA(lncRNA)AC005062.1通过调控结肠癌转移相关基因1(MACC1)的表达对结直肠癌(CRC)恶性表型的影响。方法应用GSE84983和GSE104364数据库,分析lncRNA AC005062.1在结直肠癌中的表达水平。收集医院肿瘤外科进行CRC手术治疗患者的肿瘤组织(肿瘤组)及癌旁组织(对照组),随机抽取8对,采用qPCR法检测两组中lncRNA AC005062.1的表达。运用小干扰RNA(siRNA)下调CRC细胞中lncRNA AC005062.1的表达后,采用CCK-8法检测两组细胞增殖,流式细胞方法检测两组细胞周期和凋亡,伤口划痕实验检测两组细胞迁移。利用数据库比较lncRNA AC005062.1和MACC1基因在染色上的定位,用qPCR和蛋白免疫印迹方法检测两组组织中MACC1转录水平和蛋白水平的表达;下调CRC细胞中lncRNA AC005062.1,qPCR和蛋白免疫印迹方法检测MACC1转录水平和蛋白水平的表达。结果GSE84983和GSE104364数据库分析及两组组织检测结果提示CRC中lncRNA AC005062.1呈高表达。下调lncRNA AC005062.1后,CCK-8实验结果显示,下调组HT29细胞增殖速度降低;流式细胞实验表明,下调组HT29细胞的凋亡数量增加、G_(1)期比例增加,S期和G_(2)期比例减少;Western blot实验显示下调组HT29细胞内cleaved-caspase-3和Bax表达升高,而Bcl-2表达降低;伤口划痕实验表明,下调组HT29细胞迁移率低于对照组。通过University of California Santa Cruz(UCSC)数据库分析比较显示两者在染色体上的定位很近,MACC1极可能是lncRNA AC005062.1的靶基因;检测结直肠癌两组组织中MACC1 mRNA水平,结果显示MACC1在CRC中高表达,且下调lncRNA AC005062.1在HT29细胞中的表达,可使MACC1表达降低。结论lncRNA AC005062.1可通过调控MACC1在CRC中表达,抑制结直肠癌细胞HT29增殖、周期及迁移,促进其凋亡。

Objective To explore the role and mechanism of long noncoding RNA(lncRNA)AC005062.1 in colorectal cancer(CRC)cell proliferation and apoptosis.Methods The analysis of GSE84983 and GSE 104364 was used to indicate the expression level lncRNA AC005062.1 in CRC.The tumor tissues(tumor group)and adjacent tissues(control group)of patients undergoing CRC surgery in the hospital tumor surgery department were collected.Eight pairs of tissues were randomly selected,and qPCR was used to detect the expression of lncRNA AC005062.1 in CRC tissues and paired adjacent tissues in the two groups.After using siRNA to down-regulate the expression of lncRNA AC005062.1 in CRC cells,CCK-8 assay was used to detect cell proliferation in two groups,flow cytometry was used to detect cell cycle and apoptosis in two groups,wound scratch test was used to detect cell migration in two groups,and the lncRNA was determined.The database was used to compare the localization of lncRNA AC005062.1 and MACC1 genes on staining.The expression levels of MACC1 transcription and protein levels in the control group and tumor group were detected by qPCR and Western blot,respectively.The lncRNA AC005062.1 in CRC cells was down-regulated,and the expression levels of MACC1 transcription and protein levels in the two groups of cells were detected by qPCR and Western blot,respectively.Results The database analysis of GSE84983 and GSE104364 and the detection results of the two groups indicated that the lncRNA AC005062.1 was highly expressed in CRC.After down-regulation of lncRNA AC005062.1,the results of CCK-8 experiments showed that the proliferation rate of HT29 cells in the down-regulation group decreased.Flow cytometry showed that the number of apoptosis of HT29 cells in the down-regulated group increased,the proportion of G1 phase increased,and the proportion of S phase and G2 phase decreased.Western blot experiments showed that the expressions of cleaved-caspase3 and Bax in the down-regulated HT29 cells increased,while the expression of Bcl-2 decreased.Wound scratch experiments showed that the migration rate of HT29 cells in the down-regulated group was lower than that in the control group.The analysis of UCSC to compare the location of lncRNA ac005062.1 and MACC1 on chromosomes showed that they were located very close together.MACC1 was highly expressed in CRC and down-regulated lncRNA AC005062.1 expression in HT29 cells could reduce the expression of MACC1.Conclusion lncRNA AC005062.1 can inhibit the proliferation,cycle and migration of HT29 cells,and promote its apoptosis by regulating the expression of MACC1 in CRC.